title: Nature of allosteric inhibition in glutamate racemase: discovery and characterization of a cryptic inhibitory pocket using atomistic MD simulations and pKa calculations.
abstract: Enzyme inhibition via allostery, in which the ligand binds remotely from the active site, is a poorly understood phenomenon and represents a significant challenge to structure-based drug design. Dipicolinic acid (DPA), a major component of Bacillus spores, is shown to inhibit glutamate racemase from Bacillus anthracis , a monosubstrate/monoproduct enzyme, in a novel allosteric fashion. Glutamate racemase has long been considered an important drug target for its integral role in bacterial cell wall synthesis. The DPA binding mode was predicted via multiple docking studies and validated via site-directed mutagenesis at the binding locus, while the mechanism of inhibition was elucidated with a combination of Blue Native polyacrylamide gel electrophoresis, molecular dynamics simulations, and free energy and pK(a) calculations. Inhibition by DPA not only reveals a novel cryptic binding site but also represents a form of allosteric regulation that exploits the interplay between enzyme conformational changes, fluctuations in the pK(a) values of buried residues and catalysis. The potential for future drug development is discussed.
authors.affiliation: Department of Biochemistry, University of Illinois, Urbana, Illinois 61801, USA.
authors.forename: Katie L
authors.initials: KL
authors.lastname: Whalen
authors.forename: Kenneth B
authors.initials: KB
authors.lastname: Tussey
authors.forename: Steven R
authors.initials: SR
authors.lastname: Blanke
authors.forename: M Ashley
authors.initials: MA
authors.lastname: Spies
mesh_terms: D000494:Allosteric Regulation; D000591:Amino Acid Isomerases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D055162:Biocatalysis; D007700:Kinetics; D056004:Molecular Dynamics Simulation; D016297:Mutagenesis, Site-Directed; D010848:Picolinic Acids / Q000737:chemistry; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D010848:Picolinic Acids; D011994:Recombinant Proteins; D000591:Amino Acid Isomerases; C062292:glutamate racemase; C004860:dipicolinic acid